Pancreatic adenocarcinoma is the fourth leading cause of cancer-associated mortality in the United States for both sexes, with an estimated 35,240 pancreatic cancer-related deaths in 2009. Approximately 90% of individuals present with inoperable disease at the time of diagnosis, at which point systemic therapy becomes the mainstay of their care. Currently available treatment generally consists of gemcitabine, either alone or in combination with either a second cytotoxic agent or the oral epidermal growth factor inhibitor erlotinib. Given the marginal performance status and inanition so often accompanying these patients, the rational application of non-cytotoxic targeted agents would represent an attractive alternative. We have demonstrated a negative regulatory feedback loop between two effector pathways (RAF-MEK-ERK and PI3K- AKT ) downstream of RAS, the oncogene almost ubiquitously mutated in pancreatic cancers. When breast and pancreatic cancer cell lines are treated with a MEK inhibitor, phosphorylation of the EGF receptor is markedly enhanced with resultant activation of the PI3K pathway as measured by phosphorylated AKT. Moreover, in the presence of EGF, MEK inhibitor-induced activation of AKT can be fully abolished by treatment with an EGFR inhibitor, suggesting that these cell lines have become hypersensitive to the effects of EGF. We have similarly observed at least additive benefits in vivo with dual MEK/EGFR inhibition in pancreatic tumor xenografts. In this proposal, we will therefore test the hypothesis that a dual inhibitor strategy, consisting of co-administration of a MEK inhibitor together with an EGFR inhibitor (AZD6244 plus erlotinib), will have activity in patients with gemcitabine-refractory advanced pancreatic cancer. This will be studied in the context of a phase II clinical trial that has already received approval by the Cancer Therapy Evaluation Program (CTEP). This second-line setting was selected because of the limited therapeutic options available to patients who have progressed on up-front treatment. Additionally, embedded in this clinical trial will be testing of a novel platform for collecting and analyzing circulating tumor cells (CTC), using a multidimensional cytometric scanning and imaging approach that is entirely performed on cells attached to glass slides. This platform allows us not only to quantify CTC, but also to probe them for molecular markers that may be predictive of response to dual MEK/EGFR inhibition. Further work we have planned will validate these cells as an appropriate surrogate for in situ tumor tissue by comparing them to tumor samples, whenever such material is available in our study population. Promising results from this study could justify development of a larger randomized trial comparing this combination to salvage chemotherapy, as well as evaluation in the front-line setting of advanced pancreatic cancer in conjunction with gemcitabine. Our work could also provide the rationale for similar study designs in other tumor types in which this negative MEK-EGFR-PI3K feedback loop may exist. Finally, our ability to incorporate analysis of CTC using a new technology may provide a valuable source of tumor material that can be assessed longitudinally;in the future, these cells may represent the primary source for molecular interrogation and determination of on-target treatment effects, proving useful in tailored treatment strategies for PDAC. PUBLIC HEALTH RELEVANCE: Pancreatic cancer is one of the most lethal malignancies with over 35,000 pancreatic cancer-related deaths in the United States in 2009, ranking number four in cancer- associated mortality for both sexes. This proposal uses our growing understanding of tumor biology to test a new combination of targeted agents in patients with chemotherapy-resistant, advanced pancreatic cancer, with the goal of prolonging survival in this otherwise incurable disease. Because tumor biopsy material can be so difficult to obtain in patients with advanced pancreatic cancer, we also will be studying a new technology to collect and analyze circulating tumor cells by performing a simple blood draw on patients who participate in this clinical trial.